One measuring and analytical technique in probe microscopy is scanning probe microscopy (SPM), in which a measuring probe is made to scan over a sample of measuring medium to be examined and a topography of the sample is determined by way of an gap-dependent interaction between the measuring probe and the sample. It is also possible to obtain material constants or other information on the sample. Well-known examples of this technique are the atomic force microscope (AFM) and the scanning tunneling microscope (STM). Other examples include, in particular, the scanning near field microscope (SNOM) and the scanning photone force microscope (SPhM).
In gap spectroscopy for measuring the gap-dependent interaction between the measuring probe and the sample, the measuring probe is displaced relative to the surface of the sample, for example in a vertical direction to the sample surface, and the interaction between the measuring probe and the sample is measured. Alternatively the sample may also be moved. It is also possible to provide for a relative movement between the measuring probe and the sample, in which both the measuring probe and the sample are moved. In scanning probe microscopy this gap spectroscopy for measuring the interaction between the measuring probe and the sample is used, for example, to measure forces between molecules, one molecule bonding to the measuring probe and another molecule to the sample. This is also referred to as force spectroscopy.
A component in which a base part—also known as a chip—and a thin measuring probe extension, which is also referred to as a cantilever, are formed, is generally used as measuring probe. The cantilever may be of bar-shaped design, for example, or may form a triangle with two side arms. The end of the cantilever remote from the base part (chip) may have an additional tip. In addition there are also embodiments having a plurality of extensions to the base part—so-called multi-cantilevers. Cantilevers may be composed not only primarily of silicon or silicon nitride with and without metallization, but also of plastics or polymers, such as SU-8, for example. In common usage no distinction is generally made between chip and cantilever, the entire unit instead being referred to as a cantilever. Without limiting the universality of the invention, in the following explanations reference will be made to a measuring probe. The statements will apply analogously to other forms of measuring probes in probe microscopy.
The use of both untreated and pre-treated cantilevers is known in gap spectroscopy. Using a pre-treated cantilever it is possible, in particular, to analyze specific bonds. The standard practice is to bond molecules to the measuring probe embodied as a cantilever, which together with the bonded molecule(s) then form a receptor-ligand system, for example. Also known is the practice of bonding whole cells or micro-particles to a measuring probe in the form of a cantilever and bringing this system into interaction with a sample, such as a biomaterial, for example, or with other cells. Both the coating of a pre-treated cantilever and the objects such as cells or micro-particles adhering to it will hereinafter be referred to as the probe substance.
In the probe microscopic examination the probe substance may alter and sometimes age due to contact with the sample. This may affect both the actual coating and also the state of the cells, for example in respect of the metabolism, the physiology and the activation. Furthermore it will become important, in the further use of probe microscopy in cell biology and biomedical research, to further examine the probe substance which was in contact with the sample. It would be particularly advantageous, in the handling of biological samples, if these could be used further in a germ-free or sterile and gentle manner. At present the cantilever must be removed after the measurement. To do this, the system must be taken out of the sample vessel with the surrounding fluid. This procedure in particular carries the risk that 1) the objects that have adhered are torn off by the surface tensions, 2) contamination will occur during removal of the cantilever and 3) damage to the objects will occur due to the increased time taken.
The prior art discloses how living cells as a measuring probe can be applied to an individual cantilever (cf., for example, Taubenberger A. et al.: Revealing early steps of α2β1 integrin-mediated adhesions to collagen type I using single-cell force spectroscopy, Molecular Biology of the Cell (2007) 18, 1634-1644). Here a cantilever is modified biochemically with Concavalin A, for example, so that the cell is strongly bonded to its surface once the cantilever has been brought into direct contact with the cell for a specific contact waiting time. The cell is then firmly bonded. Very large forces (typically >5 nN) are required in order to detach this cell again. Furthermore, after detaching the cell the surface is so modified that no further cell can be reattached.